Bicarbonate secretion plays an essential role in the physiological functioning of the duodenum, airway, pancreas, liver, male reproductive system, and kidney. Impairment of bicarbonate secretion, such as is manifest in the disease cystic fibrosis, can produce severe pathology in some or all of these organs. The mechanism of bicarbonate secretion is still not fully understood, despite decades of intensive study. Part of the problem is the methodology inherent in its study, which can involve intestinal, pancreatic duct, and renal tubule perfusion, and indirect methods, usually in live animals or in intact tissue. All of these methods suffer from excessive expense, cumbersome and time-consuming procedures, the use of laboratory animals which entails considerable expense and near- prohibitive regulatory requirements, and heterogeneity and lack of viability of the tissues studied. A bicarbonate-secreting in vitro system, although overcoming most of these obstacles, has not yet been developed. We propose to develop a bicarbonate secreting system based on commonly studied epithelial cell lines that have attributes similar to bicarbonate secreting intact epithelia and are thus likely to have measurable bicarbonate secretion. Our methodology includes culturing Caco-2 intestinal cells, or alternatively MDCK or other epithelial-derived cultured cells, on semi-permeable supports that can then be mounted in a perfusion chamber that enables the measurement of bicarbonate secretion, in addition to ratiometric measurement of intracellular pH, and short-circuit current across the monolayer. In this fashion, we plan to generate data that will enable us to test several key hypotheses. PUBLIC HEALTH RELEVANCE: Bicarbonate (baking soda) is secreted by the intestine in order to neutralize acid that is secreted by the stomach. Bicarbonate is also secreted by other important organs such as the pancreas, liver, lung, and male reproductive system. Disorders of bicarbonate secretion, as are found in the disease cystic fibrosis, can cause severe problems such as lung disease, pancreatitis, intestinal obstruction, and male infertility. Study of the mechanism of bicarbonate secretion has been hampered by the lack of a suitable model other than the use of live animals. We propose a cell-culture-based system suitable for the study of bicarbonate secretion that will help us and other investigators study how bicarbonate is secreted. With this knowledge, we hope to further understand why problems with bicarbonate secretion can lead to such sever diseases such as cystic fibrosis.